The research program is aimed at investigating the molecular mechanisms that make up platelet efficacy and how these can be optimally preserved during storage of a platelet transfusion product. The studies include quality monitoring of platelets collected and shipped by blood collection centers, investigating the signal transduction pathways involved in platelet activation and optimizing platelet storage under cold temperatures that minimize proliferation of bacteria. The objective is to understand how platelet activation can be minimized during storage and yet optimized in vivo when required. In addition, we are addressing current concerns about transmissible spongiform encephalopathies (TSE) and the potential for transmission of the disease by blood product transfusion. The molecular basis of TSE diseases involves conversion of prion protein(PrPc), from a normal conformation to a conformation that increases the protein's protease resistance and leads to accumulation of the protein in the central nervous system. Since blood cells express PrPc they may be able to bind and/or support proliferation the infectious conformation of prion and deliver it through transfusion to patients. The TSE diseases have a long asymptomatic phase, infected individuals could unknowingly donate blood products. Our studies investigate the physiologic role of PrPc, the role of blood cells in pathophysiology of TSE diseases in animal models and the potential for cellular blood products which express PrPc to mediate the transfer of TSE infectivity by transfusion. Evaluation of stored platelet function. The use of adenovirus, as a vector for gene therapy in a recent clinical trial with Ornithine Transcarbamylase-deficient patients, was associated with moderate to severe thrombocytopenia. In most cases the drop in platelets was reversible with a single exception where thrombocytopenia was also associated with a significant coagulopathy and ultimately a death of a study participant. The etiology of the adenovirus vector associated thrombocytopenia is not clear but may be a serious complication of gene therapy. We investigated whether the thrombocytopenia was a result of bone marrow depression by the virus (decreased production) or whether there was increased clearance of platelets from circulation (increased destruction). Adenovirus entry into cells involves two independent steps. The first step is attachment of virus to coxsackievirus adenovirus receptor, also known as CAR. The second step is internalization into the host cell and is mediated by a co-receptor, an alpha v integrin. Integrins are heterodimeric protein complexes found on many cell types; they bind adhesive proteins containing the recognition sequence arginine-glycine-aspartic acid (RDG). Examples of RGD-containing proteins include fibrinogen, fibronectin, vitronectin, and laminins. A number of integrin molecules are expressed on platelets and these include glycoprotein IIb/IIIa, the receptor for fibrinogen and vitronectin. The adenovirus could thus interact with platelets to mediate platelet clearance or to suppress their production. In a collaborative study, we followed platelet counts and circulation survival in rhesus monkeys given intravenous administration of high doses of clinical grade, replication incompetent, adenovirus. Platelet survival in circulation was determined with ex vivo biotin labeling of platelets. Reinfusion of the labeled platelets prior to administration of adenovirus and then determination of the number of labeled platelets remaining in circulation by flow cytometry. Injection of adenovirus caused a dose dependent, reversible clearance of platelets from circulation with minimal effect on platelet production. Further work will be conducted to elucidate the molecular mechanism of adenovirus induced thrombocytopenia.